1. Field of the Invention
This invention relates to a method of stimulating immune response in mammals or avian, including increasing antibody response to antigens in patients with depressed immune systems.
2. Description of Related Art
Insulin-like growth factor I (IGF-I) is a polypeptide naturally occurring in human body fluids, for example, blood and human cerebral spinal fluid. Most tissues, and especially the liver, produce IGF-I together with specific IGF-binding proteins. IGF-I production is under the dominant stimulatory influence of growth hormone (GH), and some of the IGF-I binding proteins are also increased by GH. See Tanner et al., Acta Endocrinol., 84:681-696 (1977); Uthne et al., J. Clin. Endocrinol. Metab., 39:548-554 (1974)). IGF-I has been isolated from human serum and produced recombinantly. See, e.g., EP 123,228 and 128,733.
Human growth hormone (hGH) is a single-chain polypeptide consisting of 191 amino acids (molecular weight 21,500). Disulfide bonds link positions 53 and 165 and positions 182 and 189. Niall, Nature, New Biology. 230:90 (1971). hGH is a potent anabolic agent, especially due to retention of nitrogen, phosphorus, potassium, and calcium. Treatment of hypophysectomized rats with GH can restore at least a portion of the growth rate of the rats. Moore et al., Endocrinology, 122:2920-2926 (1988). Among its most striking effects in hypopituitary (GH-deficient) subjects is accelerated linear growth of bone growth plate cartilage resulting in increased stature. Kaplan, Growth Disorders in Children and Adolescents (Springfield, Ill.: Charles C. Thomas, 1964).
It has been reported that, especially in women after menopause, GH secretion declines with age. Millard et al., Neurobiol. Aging, 229-235 (1990); Takahashi et al., Neuroendocrinology, 46:137-142 (1987). See also Rudman et al., J. Clin. Invest., 67:1361-1369 (1981) and Blackman, Endocrinology and Aging, 16:981 (1987). Moreover, a report exists that some of the manifestations of aging, including decreased lean body mass, expansion of adipose-tissue mass, and the thinning of the skin, can be reduced by GH treatment three times a week. See, e.g., Rudman et al., N. Eng. J. Med., 323:1-6 (1990) and the accompanying article in the same journal issue by Dr. Vance (pp. 52-54).
The levels of IGF-I are reported to be reduced by half in 20-month old rats compared to 6-month old rats. Takahashi and Meiters, Proc. Soc. Exp. Biol. Med., 186:229-233 (1987). See also Florini and Roberts, J. Gerontol., 35:23-30 (1980); Florini et et al., Mech. Aging Dev., 15:165-176 (1981); Chatelain et al., Pediatrie, 44:303-308 (1989); Florini et al., J. Gerontol., 40:2-7 (1985); Hall and Sara, Clinics in Endocrin, and Metab., 15:629 (1984); Baxter, Advances in Clinical Chemistry, 25:49 (1986); Clemmons and Underwood, Clinics in Endocrin. and Metab., 28:629 (1986); Hintz, Advances in Pediatrics, 28:293 (Year Book Medical Publishers, Inc., 1981); Johanson and Blizzard, The Johns Hopkins Medical Journal, 149:115-117 (1981), the latter five references describing low IGF-I levels in aged men. The Hintz, Clemmons and Underwood, and Baxter references are general reviews on IGF-I.
Furthermore, it was found that among human diploid fibroblasts capable of cycling in aging cultures in vitro, there were few changes in the regulation of the growth fraction by platelet-derived growth factor (PDGF) and epidermal growth factor (EGF), but a greatly increased dependence on IGF-I for regulation of the rate of entry into S phase. Chen and Rabinovitch, J. Cell. Physiol., 144:18-25 (1990). The authors conclude that the slower growth of the dividing population of cells in aging cultures may be related to a requirement for IGF-I at levels that are greatly above those usually supplied. This may be due to overproduction of the IGF-I binding protein, IGFBP-3, and, therefore, a reduction in IGF-I availability to its receptor. Goldstein et al., "Cellular and Molecular Applications to Biology of Aging", AFCR Meeting abstract, Seattle, May 4-5, 1991.
Various biological activities of IGF-I in other than aged mammals have been identified. For example, IGF-I is reported to lower blood glucose levels in humans. Guler et al , N. Engl. J. Med., 317:137-140 (1987). Additionally, IGF-I promotes growth in several metabolic conditions characterized by low IGF-I levels, such as hypophysectomized rats [Skottner et al., J. Endocr., 112:123-132 (1987)], diabetic rats [Scheiwiller et at., Nature, 323:169-171 (1986)], and dwarf rats.[Skottner et al., Endocrinology, 124:2519-2526 (1989)]. The kidney weight of hypophysectomized rats increases substantially upon prolonged infusions of IGF-I subcutaneously. Guler et al., Proceedings of the 1st European Congress of Endocrinology, 103: abstract 12-390 (Copenhagen, 1987). The kidneys of Snell dwarf mice and dwarf rats behaved similarly. van Buul-Offers eg al., Pediatr. Res., 20:825-827 (1986); Skottner et al. Endocrinology, supra. An additional use for IGF-I is to improve glomerular filtration and renal plasma flow. Guler et al., Proc. Natl. Acad. Sci. USA, 86: 2868-2872 (1989). The anabolic effect of IGF-I in rapidly growing neonatal rats was demonstrated in vivo. Philipps et al., Pediatric Res., 23:298 (1988). In underfed, stressed, ill, or diseased animals, IGF-I levels are well known to be depressed.
GH and IGF-I have been linked with immunoregulatory properties. The immune response results from interaction of antigens (foreign or non-self moieties) with host cells (lymphocytes) bearing specific receptors on the surface membrane for these antigens. Lymphocytes are grouped into two major classes, T-cells and B-cells.
T-cells originate from the thymus where they mature and differentiate from bone-marrow-derived cells. The mature T-cells leave the thymus gland to continuously circulate from blood to lymph nodes and spleen and back to blood. T-cells are further subdivided into three major subsets: T-helper cells, T-suppressor cells, and T-cytolytic cells. T-helper cells "help" other cells: B-cells to secrete antibody, cytotoxic cells to become functional, and macrophages to become activated. This population of T-cells bears the CD.sub.4 surface marker that is used to identify this subset in tissue and blood.
T-cytolytic cells are responsible for killing target cells such as virally infected cells, tumor cells, and allografts. Suppressor T-cells act to limit and terminate the immune response. The cytolytic and suppressor T-cell populations are identified by the CD.sub.8 surface marker.
The B-cells, or antibody-forming cells, also derive from immature precursors found in the bone marrow. When mature, the B-cells migrate to all lymphoid organs except the thymus. B-cells interact with antigens by way of antibody molecules bound to their plasma membranes that act as receptor proteins. This surface immunoglobulin is used as a marker to identify B-cells in tissue and blood. Following interaction with antigen and T-helper cells, the B-cells differentiate into antibody-forming cells called plasma cells. These plasma cells secrete antibody into the extracellular matrix. The antibody diffuses into capillaries and circulates via normal blood flow. Thus, the serum immunoqlobulin level reflects the cellular dynamics of the immune response.
In many states, children are required to be immunized routinely against such diseases as diphtheria, pertussis, and typhoid (DPT), as well as measles, tetanus, mumps, polio, and rubella, by administering vaccines. The B-cell reaction to vaccine is the production of appropriate immunoglobulins, which are intended to confer immunity against the disease. Generally, a particular B-cell will be differentiated to produce one particular type of antibody, and such production is caused by the presence in the body of one particular type of antigen. Hence, when an animal or person has been exposed to a number of different antigens, the animal or human will have a number of different B-cells that can produce its particular immunoglobulins when the appropriate antigen is present.
In some situations, the immune response to antigen is insufficient to confer immunity. That is, a quantity of immunoglobulins is generated (or a number of B-cells are potentiated) that is insufficient to confer effective immunity.
It has been known since 1967 that a connection exists between the anterior pituitary and the immune system, and specifically with GH. Two groups of investigators concluded from their studies that GH controls the growth of lymphoid tissue. Pierpaoli and Sorkin, Nature, 215:834 (1967); Baroni, Experientia, 23:282 (1967). Subsequently, immunologic function was restored in the pituitary dwarf mouse by a combination of bovine somatotropic hormone and thyroxin. Baroni et al., Immunol., 17:303-314 (1969).
In a sex-linked dwarf chicken strain, bovine GH treatment resulted in enhanced antibody responses and bursal growth while thyroxine treatment stimulated thymus growth. Marsh et al., Proc. Soc. Exp. Biol. Med., 175:351-360 (1984). However, neither treatment altered immune function in the autosomal dwarf chicken. Bovine GH therapy alone partially restored immunologic function in immunodeficient Weimaraner dogs. Roth et al., Ann. J. Vet. Res., 45:1151-1155 (1984).
Mice with hereditary GH deficiency develop an impairment of the immune system associated with thymic atrophy, immunodeficiency, and wasting, resulting in a shortened life expectancy. Frabris et al., Clin. Exp. Immunol., 9:209-225 (1971). It has been shown that an age-associated decline in the plasma concentration of thymulin (a thymic hormone) occurs and that plasma thymulin concentration increases in bGH-treated middle-aged and old dogs. Goff et al., Clin. Exp. Immunol., 68:580-587 (1987). The authors suggest that exogenous GH may be useful for restoring some immune functions in aged individuals. Further, administration of hGH to C.sub.57 /B1/6J mice was found to reverse the inhibitory effect of prednisolone on thymus and spleen cellularity and on natural killer activity; administration of hGH without prednisolone had no effect, although at higher doses it induced a decrease of thymic parameters and natural killer activity with no effect on spleen cellularity, and relative weights. Franco et al., Acta Endocrinologica, 123:339-344 (1990).
It has also been shown that GH induces T-cell proliferation in the thymus. Murphy et al., FASEB Meeting Abstract, Atlanta, April 1991; Durum et al., FASEB Meeting Abstract, Atlanta, April 1991. For recent reviews on the immune effects of GH, see Kelley, "Growth Hormone in Immunobiology," in Psychoneuroimmunology II, 2nd Ed., B. Ader et al., eds., Acad. Press 1990, and Ammann, "Growth Hormone and Immunity," in Human Growth Hormone--Progress and Challenges, L. Underwood, ed., Marcel Dekker, Inc., New York, (1988), pp. 243-253; Weigent and Blalock, Prog. NeuroEndocrinImmunology, 231-241 (1990). It has been reported that the activity of all major immune cell types, including T-cells, B-cells, natural killer (NK) cells and macrophages, can be altered by GH. Kelly, Biochem. Pharmacol., 38:705 (1989).
One report states that locally generated IGF-I mediates GH action on T-lymphocytes through the type I IGF receptor. Geffner et al., J. Clin. Endocrin, and Metab., 71:464 (1990 ). Also, Franco et al., on p. 343, speculate that some of the effects of hGH on the immune system occur via IGF-I. Timsit et al., 73rd Annual Meeting, Endocrine Society, June 19-22, 1991, abstract 1296, reports hGH and IGF I stimulate thymic hormone function.
There have been data published documenting the ability of cells of the immune system to produce IGF-I-like molecules. These include activated alveolar macrophages [Rom et al., J. Clin. Invest., 82:1685 (1988)], human B-lymphocytes transformed with Epstein-Barr virus [Merimee et al., J. Clin. Endocrin. Metab., 69:978 (1989)], spleen and thymus tissues through detection of mRNA for IGF-I [Murphy et al., Endocrinology, 120:1279 (1987)], and normal T-cells [Geffner et al., supra].
Data have also been presented suggesting that IGF-I produced locally in tissues such as the thymus or inflammatory sites might affect the growth and function of IGF-I-receptor-bearing T-lymphocytes. Tapson et al., J. Clin. Invest., 82:950-957 (1988).
A statistically significant increase in thymus and spleen weight of hypophysectomized rats infused for 18 days with IGF-I was observed as compared to control or treatment with GH. Froesch et al., in Growth Hormone Basic and Clinical Aspects. eds. O. Isaksson et al., p. 321-326 (1987). Also reported was an increased thymic tissue in young GH-deficient rats treated with IGF-I [Guler et al., Proc. Natl. Acad. Sci. USA, 85:4889-4893 (1988)] and an increase in the spleen of dwarf rats [Skottner et al., Endocrinology, supra]. Others have shown repopulation of the atrophied thymus in diabetic rats using either IGF-I or insulin; however, when the rats were immunized with bovine serum albumin (BSA) and boosted, serum anti-BSA antibodies showed no effect of insulin or IGF-I on the antibody response despite large effects on thymic and splenic size. Binz et al., Proc. Natl. Acad. Sci. (USA). 87:3690-3694 (1990). IGF-I was reported to stimulate lymphocyte proliferation (Johnson et al., Endocrine Society 73rd Annual Meeting, abstract 1073, June 19-22, 1991).
Furthermore, IGF-I was found to repopulate the bone marrow cavity with hematopoietic cells [Froesch et al., supra], stimulate erythropoiesis in hypophysectomized rats [Kurtz et al., Proc. Natl. Acad. Sci. (USA). 85:7825-7829 (1988)], and enhance the maturation of morphologically recognizable granulocytic and erythroid progenitors in suspension cultures of marrow cells. Merchav et al., J. Clin. Invest., 81:791 (1988).
At nanomolar concentrations, IGF-I is a growth-promoting factor for lymphocytes. Schimpff et al., Acta Endocrinol., 102:21-25 21-25 (1983). B-cells, but not T-cells, have recently be to possess receptors for IGF-I. Stuart et al., J. Clinical Endo. and Met., 72:1117-1122 (1991). Also, IGF-I, as a chemotactic for resting and activated T-cells, stimulates an increase in thymidine incorporation into resting and activated T-cells. Normal T-cell lines show augmentation of basal colony formation in response to IGF-I. Geffner et al., supra. It is also stated on p. 955 of Tapson et al., J. Clin. Invest., 82:950-957 (1988) that IGF-I produced locally in tissues such as the thymus or inflammatory sites might affect the growth and function of IGF-I receptor-bearing T lymphocytes. However, IGF-I is reported to suppress in a dose-dependent manner IL-2-induced proliferative responses and in vitro antibody responses of splenocytes. Hunt and Eardley, J. Immunol., 136:3994-3999 (1986).
There is a need in the art to supply a reagent that will stimulate the immune system of a mammal or avian, whether the immune response is cell-mediated or antibody-mediated. There is a particular need for a reagent that will boost the antibody response of patients with compromised immune systems to antigens to which they are exposed. In view of the controversy in the art surrounding IGF-I, it is unclear what its effects would be in increasing immune function, as opposed to merely increasing size of organs involved in immune function such as the thymus and spleen, or in increasing the activity of T- or B-cells in vitro or in vivo.
It is therefore an object of the present invention to stimulate the immune response of a mammal or avian.
It is a particular object to increase production of immunoglobulins by increasing the number of immunoglobulin-producing cells and/or by increasing the amount of immunoglobulin produced by the individual immunoglobulin-producing cells in response to the predetermined immunogen.
It is a more particular object to increase antibody responses in patients with severely hampered immune systems, such as patients who receive bone marrow transplants or in AIDS patients.
These and other objects will be apparent to those of ordinary skill in the art.